I have been extracting protein from various seed meals but am having trouble visualizing them on my gel. I have tried two different types of gels, Tris-Acetate and Tricine, but so far neither have shown any bands. Should I be staining the gels or using UV to visualize bands? I was under the assumption that the loading buffer would stain the proteins.
The loading buffer will not stain your sample. Usually, protein loading dye/buffer has SDS that helps to denature the proteins and to make them negatively charged. Also, it has glycerol to ensure that your protein sample won't float away, as glycerol has high density. It also has thiol agents that work with SDS for successful protein linearization. The bromphenol blue, or the colour dye in the buffer helps in visualizing your sample in the well and tracking its progress through the gel later. For visualization, you should stain them. Hope this helps.
Did you quantified you protein before load in in the gel?
Hoh did you preparare the sample before loading into the gel?
Because SDS page do not have an high sensitivity and you need to load ugs of proteins.
best regards
Manuele
Hi Kyle,
As previously said your sample buffer is not a way to stain your proteins but just to condition them for proper migration and follow the whole migration front. To see your proteins after migration you can stain your gel with Coomassie blue or silver nitrate if the former one is not sensitive enough. If you still do not see anything then you need to increase the amount of material loaded on your gel.
Good luck.
Hi Kyle,
The loading buffer will not stain your sample, but you can stain your gel with Coomassie blue or another stain after protein migration. I suggest watching this helpful video on SDS-PAGE! https://www.youtube.com/watch?v=mabx-U5X9J8
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